邻苯二甲酸酯及双酚A或氯化镉对睾丸巨噬细胞分泌白细胞介素-10的影响

Effects of joint exposure to DEHP,BPA and Cd Cl2on IL-10 produced by testicle macrophage

目的研究邻苯二甲酸二(2-乙基)己酯(DEHP)及双酚A(BPA)或氯化镉对大鼠睾丸巨噬细胞(TM)分泌细胞因子IL-10的影响。方法原代提取大鼠TM细胞,以50μg/ml脂多糖(LPS)作为激活剂,分别设1、10、100μmol/L DEHP、BPA和1、10、50μmol/L氯化镉单独染毒组以及0.5μmol/L DEHP+0.5μmol/L BPA、5μmol/L DEHP+5μmol/L BPA、50μmol/L DEHP+50μmol/L BPA、0.5μmol/L DEHP+0.5μmol/L氯化镉、5μmol/L DEHP+5μmol/L氯化镉、50μmol/L DEHP+25μmol/L氯化镉联合染毒组。作用细胞24 h后,采用ELISA及细胞因子抗体芯片方法检测TM细胞上清液中细胞因子白细胞介素-10(IL-10)的浓度。结果与LPS组比较,1μmol/L氯化镉、10μmol/L BPA或100μmol/L DEHP单独染毒组的IL-10蛋白均较低,差异有统计学意义(P<0.05);且随着剂量的增加抑制作用逐渐增强。无论与LPS组还是与1μmol/L BPA单独染毒组比较,0.5μmol/L DEHP+0.5μmol/L BPA混合染毒组的IL-10蛋白降低,差异有统计学意义(P<0.05)。抗体芯片结果显示50μmol/L DEHP+50μmol/L BPA混合染毒组的荧光值是100μmol/L DEHP单独染毒组的76.9%,是100μmol/L BPA单独染毒组的115.0%。无论与LPS组还是100μmol/L DEHP单独染毒组比较,50μmol/L DEHP+25μmol/L氯化镉混合染毒组TM细胞上清液中IL-10蛋白浓度均较低,差异有统计学意义(P<0.05)。结论 DEHP及BPA或氯化镉单独作用能明显抑制TM细胞分泌IL-10蛋白。低剂量(0.5μmol/L)DEHP和BPA的联合作用为协同作用,中、高剂量(5μmol/L、50μmol/L)的联合作用为拮抗作用。而DEHP和氯化镉对IL-10蛋白的交互作用为协同作用。

Objective To investigate the effect of phthalate（2-ethylhexyl） ester（DEHP）,bisphenol A（BPA） and cadmium chloride（Cd Cl2） on interleukin-10（IL-10） produced by primaryly cultured rat＇s testicle macrophage（TM） and the combined effects. Methods Primaryly cultured TM were stimulated with 50 μg/ml lipopolysaccharide（LPS） as activating agent and exposed to the different doses of DEHP（1,10 and 100 μmol/L）,BPA（1,10,100 μmol/L） and Cd Cl2（1,10,50 μmol/L） alone or joint treatment（0.5 μmol/L DEHP＋0.5 μmol/L BPA, 5 μmol/L DEHP＋5 μmol/L BPA, 50 μmol/L DEHP＋50 μmol/L BPA, 0.5μmol/L DEHP＋0.5 μmol/L Cd Cl2,5 μmol/L DEHP＋5 μmol/L Cd Cl2, 50 μmol/L DEHP＋25 μmol/L Cd Cl2）. Interleukin-10（IL-10） in the culture supernatants was determined by cytokines antibody microarrays and ELISA. Results Compared with LPS group,the levels of IL-10 protein in the culture supernatant decreased obviously in 1 μmol/L Cd Cl2,10 μmol/L BPA and 100μmol/L DEHP exposure groups. The inhibitory effect enhanced gradually with the increased doses. Compared with LPS or 1μmol/L BPA groups,the levels of IL-10 were inhibited obviously in 0.5 μmol/L DEHP ＋0.5 μmol/L BPA group. The fluorescence value of 50 μmol/L DEHP＋50 μmol/L BPA group was 76.9% of that in 100 μmol/L DEHP group and 115.0% in100 μmol/L BPA group showed by Rat Cytokine Antibody Microarray. Compared with LPS or 100 μmol/L DEHP groups,the levels of IL-10 obviously decreased in 50 μmol/L DEHP＋25 μmol/L Cd Cl2 group. Conclusion DEHP,BPA and Cd Cl2 single exposure may disturb IL-10 protein produced by TM cells. The combined exposure of DEHP and BPA may present synergistic effect in low dose（0.5 μmol/L） and antagonistic action in moderate（5 μmol/L） or high doses（50 μmol/L）. However, the joint exposure of DEHP and Cd Cl2 may have synergistic effect.